CHX and a Face Shield Cannot Prevent Contamination of Surgical Masks.

Background: Bacterial contamination on surgical masks puts a threat to medical staff and patients. The aim of the study was to investigate its contamination during dental treatments, wearing a face shield and performing a pre-procedural mouth rinsing with chlorhexidine (CHX). Methods: In this prospective, randomized study, 306 treatments were included, 141 single-tooth (restorations) and 165 total dentition treatments (preventive or periodontal supportive ultrasonic application). A total of three groups (each: n = 102) were formed: participants rinsed for 60 s with 0.1 % CHX or with water before treatment, and, for control, a non-rinsing group was included. In view of the COVID-19 pandemic, a face shield covering the surgical mask enhanced personal protective equipment. After treatment, masks were imprinted on agar plates and incubated at 35°C for 48 h. Bacteria were classified by phenotypic characteristics, biochemical assay methods, and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS). Colonies (CFU) were counted and mean values were compared (Kruskal-Wallis-, U test, p < 0.05). Results: Chlorhexidine led to a statistically significant reduction of bacterial contamination of the surgical mask (mean: 24 CFU) in comparison with water (mean: 47 CFU) and non-rinsing (mean: 80 CFU). Furthermore, rinsing with water reduced CFU significantly in comparison with the non-rinsing group. There were no significant differences between single or total dentition treatments. Streptococcus spp., Staphylococcus spp., Micrococcus spp., and Bacillus spp. dominated, representing the oral and cutaneous flora. Conclusion: A pre-procedural mouth rinse is useful to reduce the bacterial load of the surgical mask. However, contamination cannot be prevented completely, even by applying a face shield. In particular, during pandemic, it is important to consider that these additional protective measures are not able to completely avoid the transmission of pathogens bearing aerosols to the facial region. If antiseptic rinsing solutions are not available, rinsing with water is also useful.

Ribozyme Assays to Quantify the Capping Efficiency of In Vitro-Transcribed mRNA.

The presence of the cap structure on the 5'-end of in vitro-transcribed (IVT) mRNA determines its translation and stability, underpinning its use in therapeutics. Both enzymatic and co-transcriptional capping may lead to incomplete positioning of the cap on newly synthesized RNA molecules. IVT mRNAs are rapidly emerging as novel biologics, including recent vaccines against COVID-19 and vaccine candidates against other infectious diseases, as well as for cancer immunotherapies and protein replacement therapies. Quality control methods necessary for the preclinical and clinical stages of development of these therapeutics are under ongoing development. Here, we described a method to assess the presence of the cap structure of IVT mRNAs. We designed a set of ribozyme assays to specifically cleave IVT mRNAs at a unique position and release 5'-end capped or uncapped cleavage products up to 30 nt long. We purified these products using silica-based columns and visualized/quantified them using denaturing polyacrylamide gel electrophoresis (PAGE) or liquid chromatography and mass spectrometry (LC-MS). Using this technology, we determined the capping efficiencies of IVT mRNAs with different features, which include: Different cap structures, diverse 5' untranslated regions, different nucleoside modifications, and diverse lengths. Taken together, the ribozyme cleavage assays we developed are fast and reliable for the analysis of capping efficiency for research and development purposes, as well as a general quality control for mRNA-based therapeutics.

BNT162b2 vaccine induces neutralizing antibodies and poly-specific T cells in humans.

BNT162b2, a nucleoside-modified mRNA formulated in lipid nanoparticles that encodes the SARS-CoV-2 spike glycoprotein (S) stabilized in its prefusion conformation, has demonstrated 95% efficacy in preventing COVID-191. Here we extend a previous phase-I/II trial report2 by presenting data on the immune response induced by BNT162b2 prime-boost vaccination from an additional phase-I/II trial in healthy adults (18-55 years old). BNT162b2 elicited strong antibody responses: at one week after the boost, SARS-CoV-2 serum geometric mean 50% neutralizing titres were up to 3.3-fold above those observed in samples from individuals who had recovered from COVID-19. Sera elicited by BNT162b2 neutralized 22 pseudoviruses bearing the S of different SARS-CoV-2 variants. Most participants had a strong response of IFNγ+ or IL-2+ CD8+ and CD4+ T helper type 1 cells, which was detectable throughout the full observation period of nine weeks following the boost. Using peptide-MHC multimer technology, we identified several BNT162b2-induced epitopes that were presented by frequent MHC alleles and conserved in mutant strains. One week after the boost, epitope-specific CD8+ T cells of the early-differentiated effector-memory phenotype comprised 0.02-2.92% of total circulating CD8+ T cells and were detectable (0.01-0.28%) eight weeks later. In summary, BNT162b2 elicits an adaptive humoral and poly-specific cellular immune response against epitopes that are conserved in a broad range of variants, at well-tolerated doses.